Thereisacriticalneedforimprovedhumantissuemodelstostudyinfectiousdiseases.Animalmodelsoften failtoreproducehumanphysiology,andaresimilarlypoorpredictorsofdrugefficacywhentranslatedto humans.ThisproposaldescribestheMITCenterforHumanTissueModelsforInfectiousDiseases (MIT.HTMID),whichwillfocusontwodimensionalhumanneuralcellsandthreedimensionalhumancerebral organoidstostudyvirusinfections.TheprojectissitedentirelyatTheMassachusettsInstituteofTechnology. Thethreeinvestigatorsare:LeeGehrke(VirologyandInfectiousDiseases),RudolfJaenisch(Tissue Engineering,CellBiology,andStemCells)andDavidSabatini(GeneticsandScreeningTechnologies).The CenterwillalsoincludethreeCores?thatis,Administrative,Virology,andHumanCellsandTissues.The themesofthetwointerrelatedResearchProjectsofthisU19proposalare:?Project1:?Humantissuemodelsto studyinfectiousdiseases:Human2Dand3Dneuralculturesforstudyingvirustropismandinfection phenotypes,and?Project2:?Useof2Dculturesand3Dorganoidstoidentifycandidateantiviralcompounds?to usegeneticapproachestoidentifyandvalidatehostgenesthatpromoteorprotectagainstflavivirusinfection?. Theexperimentaluseoforganoidsissignificantbecausethethreedimensionalarchitectureanddifferentiation fromembryonicstem(ES)cellsandinducedpluripotentstem(iPS)cellsprovidenearphysiologicalfunctionsin tissueorganization,tissuerenewal,andresponsestopathogeninfections.Indeed,humanorganoidshave beengeneratedforawiderangeoftissuesandusesinstudyingdevelopmentanddiseases,includingvirus infections.Theprojectwillcomparetheinfectionsoffivedifferentneuralormicroglialcelltypes(neuronal progenitors,neurons,oligodendrocytes,astrocytes,microglia)withthreeflaviviruses(ZikaVirus,WestNile Virus,orDenguevirus).TheresearchgoalsofMIT.HTMIDaddresstheZikavirusglobalhealthcrisis,toward understandinghowrelatedflavivirusescancauseverydifferentdiseases,includingmicrocephalyandGuillain BarreSyndrome.Thevirusworkwillbeextendedbeyondflavivirusestoincludeotherneurotropicviruses?that is,pseudotypedvesicularstomatitisviruses(VSV)thatcarrytheenvelopesofselectagentencephaliticviruses (EasternEquineEncephalitis,WesternEquineEncephalitis,andVenezuelanEquineEncephalitis).Wewill usecellandmolecularmethodstodefineandcomparetheinfectionphenotypesofthecellsandviruses. TissueengineeringandgeneticswillbecombinedbyperformingCRISPRCas9screenstoidentifygenesthat regulateorareregulatedbyvirusinfection,andthengenerating?knockout?organoidstotestfunctionina threedimensionaltissue.Theorganoidhumantissuemodelwillalsobeusedtoevaluateanumberofantiviral compoundstovalidateitspotentialuseasadrugtestingplatform.Ifsuccessful,thisCenterwillyieldsignificant newdataandbeanintegralcomponentofanetworkofHumanTissueModelsforInfectiousDiseases.